About this Author

College chemistry, 1983

The 2002 Model

After 10 years of blogging. . .

Derek Lowe, an Arkansan by birth, got his BA from Hendrix College and his PhD in organic chemistry from Duke before spending time in Germany on a Humboldt Fellowship on his post-doc. He's worked for several major pharmaceutical companies since 1989 on drug discovery projects against schizophrenia, Alzheimer's, diabetes, osteoporosis and other diseases.
To contact Derek email him directly: derekb.lowe@gmail.com
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April 18, 2012

Buckyballs Prolong Life? Really?

Posted by Derek

I'm really, really not sure what to make of this paper (PDF). It's from a team that was studying the long-term toxicology of C60 (fullerene, "buckyballs") by giving them to rats as a solution in olive oil. The control groups were water and olive oil without C60. The compound has already been shown to have no noticeable short-term toxic effects, so they probably didn't expect anything dramatic in the lower-dose long-term mode.

Wrong. What they found was that the fullerene/olive oil group had their life spans extended by some 90%, which would make this mixture perhaps the most efficacious life-extended treatment ever seen in a rodent model. This is a very odd and interesting result.

There's nothing bizarre about the pharmacokinetics, anyway. A reasonable amount of the C60 is absorbed after an oral dose (they did both oral gavage and intraperitoneal dosing), with a time course consistent with the very high lipophilicity of the compound. Distribution is still being worked out, but a lot of any given dose ends up in the liver and spleen (although it doesn't accumulate with successive q.d. dosing), with detectable amounts even crossing the blood-brain barrier. It has a long half life, consistent with enterohepatic recirculation and elimination through the bile (no C60 was found in the urine).

The most likely mechanism for the life-extension effects is through oxidative stress and free radical scavenging. There have been several reports of C60 as an antioxidant, although there have also been reports that it can be cytotoxic via lipid peroxidation. (One difference was that that report was with aggregates of C60 in water, versus soluble C60 in oil, but there are other reports that hydrated C60 does the opposite: there's clearly a lot that hasn't been cleared up here). In this study, even at very low doses, C60 appears to protect rodents against carbon tetrachloride-induced liver damage, for example, which is known to involve a radical process. Significantly, it does so while showing protection against glutathione depletion, which also suggests that it's directly scavenging reactive intermediates.

These are reasonable (but unproven) hypotheses, and I very much look forward to seeing this work followed up to see some more light shed on them. The whole life-extension result needs to be confirmed as well, and in other species. I congratulate the authors of this work, though, for giving me the most number of raised eyebrows I've had while reading a scientific paper in quite some time.

First, can we get something straight on the use of the term "scavenging"? The cumulative dose over 7 months in this study was ~200mg or 275 micromoles. This is not enough for it to be working in a scavenging mode (like ascorbate or glutathione) where the product is disposed of or recyclesd. If it is working via affecting ROS levels at all, it has to be acting in a catalytic mode, or it is being recycled by other antioxidant systems. Maybe it's increasing the efficiency of the tocopherol/ascorbate shuttle, but it can't simply be sopping up radicals like a sponge.

Second, I find it VERY odd that they started dosing at 10 months (just a few months before the control rats started dying), and kept the therapy up for only 7 months, but the oldest lived rats went on for over 5 years. Really? In humans this would translate to taking C60 for most of your 30s, none before, none after, and living to 120. Something's not right.

My other suspicion comes from the timing... the oldest lived animals were 66 months (5.5 years), and the paper was submitted in Jan 2012. Thus, the studies were begun in July 2006. Many of the papers they cite regarding the rationale for the dosing regimen were published AFTER the study had already begun. The paper providing the rationale for stopping at 7 months (due to accumulation of c60 in the liver) was not published until 2010, more than 3 years after they'd already stopped dosing! You can't cite something as rationale, when it wasn't even published when you were designing the study.

Isn't it odd that the estimated median lifespans (EMLs) for the olive oil and C60-olive oil treated rats are a lot shorter than the time it took for the first rat to die (compare Figure 3 to the text on 4, top left column)? Maybe it's because I don't understand non-parametric Kaplan-Meier analysis, but this seems like a non-sensical result. Can anyone clarify how this can happen?

That doesn't change the qualitative conclusion that adding C60 does something, but it does change the quantitative evaluation of the relative effects of olive oil and C60, which in turn affects determination of the synergy between C60 and olive oil and that could be significant.

@9:
You don't want rats to live longer. The nice thing about pet rats is they die off just about the time the kid you bought it for loses interest in it and just before the parent has to start taking care of it.

I was once with a company which had begun to market rodents which were GMd to be prone to developing tumors. I suggested that we could make a killing in the home market by selling these as pets guaranteed to die after six months. Marketing did not seize upon this idea.

@8 I always thought that median time to death was the time it took for 50% of the animals to die. Eyeballing figure 3, this is 26, 42 and 63 months for water, oil, oil + C60 respectively. No rats in the C60 group died before 60 months, so I don't see how median survival for this group can be the stated 42 months unless I'm missing something (always a fair assumption).

bacillus, #14 OK, so if I'm clueless, at least I'm not alone. Maybe the shorter-than-we-would've-expected median lifespans have something to do with non-parametric parameters in the non-parametric Kaplan-Meier analysis, eh? :)

@16. It looks like controls die at around 16,26,29,32,35,38 months; oil at 36,38,40,43,58,58 months; C60 at 60,61,61,63,64,64 months which when plugged into Prizm statistical analysis gives median survival of 30, 40, 62, months. However, I am not a statistician.

Overall, the study seems to be a bit fishy to me. A lifespan of 5.5 years in a laboratory rat seems rather astonishing, considering that we toxicologists rarely see rats reaching the age of 3 or 4 years if left to themselves. And anybody else notice that they do not give information on the rat strain used, or the diet given? It is well known that dietary calory intake restrictions leed to an increase in lifespan. Equally interesting would be to know how the authors might explain the observed effect after weekly dosing when the half life of the compound is 9 hours (table 1). And heck, if their results hold true, why on earth did they publish in Biomaterials and not in a respectable biomedical or toxicological journal? I'd die to have a look at the peer reviewers' and editor's comments....

They only have SIX animals per cohort! That's way too low to get valuable information on lifespan. If they're serious about this they'll repeat it with at least 40 animals per group of each sex. It's expensive and time consuming, but the data is pretty useless otherwise.

Look at the images on page 7 of the paper. Specifically the top ones for the GAip versus GAop liver biopsy's. Anyone else see what I see? Sloppy at best, fraud at worst. I figure that people would have learned their lessons after the Mikovits XMRV photoshopping incident.

I brought this up with my research group (organic electronics) and the boss was skeptical that they'd even be able to dissolve 0.8mg/ml fullerene in olive oil - it isn't known to have any good aliphatic solvents (but it obviously would have been hard to get results feeding the rats CS2 or toluene).

I agree as flatlander has noted that the images in the top two panels are the same. Also, the images are blurry and poorly focused. Can't hardly see much detail. Changes are prominent enough that things can be seen. The "C60 crystals" in Figure 2 are not "specific" as described by the authors. Looks like it could be hemosiderin (normal finding in rat spleen) and special stains are needed to confirm.

I am highly skeptical, and it looks like more of a sales pitch for Tunisian olive oil than BB, the way parts of it is written. I find it hard to believe that olive oil alone will extend a rat's life as long as they claim. I also wouldn't have started the chronic study with rats 10 months old.

Note in table 2 the organ weights. Liver weights of ~5, 7, 7 and 10 across the 4 data points? you need a potent PPAR alpha to get doubling of liver weights, and I'm not sure you can do that in just one week.

Haven't waded through the rest of the details, but I suspect that there was a journal selection bias by the authors to get this paper published. I also agree that it should've gone to a good tox or tox path journal instead (but if it had was likely turned away).

The ageing field is, to put it mildly, problematic. Too many snake oil salesmen and too little credible science. Death of a laboratory animal is not a great endpoint-very often the mode of death is not really understood at the mechanistic level.
Last week, I sat though a truly awful fly talk, which culminated in the hapless presenter admitting that she didn't know why the flies died, but only that the rate of death was slightly lower...
Allied to Big Pharma's search for something to sell to the healthy rich, and it's a recipe for confusion/Sirtris.

@37, bacillus. You raise another good point. The article gives the impression that this work was a chronic tox study that just happened to observe something astonishing. But who plans to run a chronic tox study for that long? It's very unusual. Normally, you might run it for a year, then sacrifice the surviving animals for histopathology. It's almost as if they planned the experiment with an end result in mind...

I race bicycles in the Master's 50+ category for a team sponsored by Colavita. At the start of one of the races, the race marshall looks at me and says, "Hey I might need to see some ID from you". I shot back while pointing to the logo on my sleeve, "I've got two words for ya buddy, Olive Oil"!